Targeting of nanoparticles to cell adhesion molecules for potential immune therapy

Abstract

Cell adhesion molecules including leukocyte function associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) play an important role in regulating inflammatory responses. For circulating leukocytes to enter inflamed tissue or peripheral lymphoid organs, the cells must adhere to and transmigrate between endothelial cells lining blood vessel wall by binding of LFA-1 on leukocytes to ICAM-1 on endothelial cells. In addition, interaction of LFA-1 expressed on T cells and ICAM-1 expressed on antigen presenting cells (APCs) is crucial for immunological synapse formation and hence T cell activation. Clustering of LFA-1 and ICAM-1 by multivalent ligands increases binding avidity of these cell adhesion molecules. In this thesis, multivalent ICAM-1 or LFA-1 ligands were conjugated to the surface of polymeric nanoparticles (NPs) to target the clustering of these receptors and increase the avidity of binding to ICAM-1 or LFA-1. Polymeric nanoparticles possess some advantages over other multivalent ligands since drugs can be protected and released. This delivery system permits modification of the nanoparticle surface without compromising the activity of the drug carried. In chapter 2 of this thesis, a peptide ligand targeting ICAM-1 (cLABL) was conjugated to poly (DL-lactic-co-glycolic acid) (PLGA) nanoparticles. The cellular uptake of cLABL conjugated NPs (cLABL-NPs) by lung carcinoma epithelial cells upregulating ICAM-1 was significantly more rapid than control NPs. The specificity of ICAM-1 mediated internalization was confirmed by blocking the uptake of cLABL-NPs to ICAM-1 using free cLABL peptide. Cell studies suggested that cLABL-NPs targeted encapsulated doxorubicin to ICAM-1 expressing cells and provided sustained release of doxorubicin. In chapter 3, a peptide ligand targeting LFA-1 (cIBR) was conjugated to PLGA NPs to specifically target T cells expressing LFA-1. The specificity of NPs targeting LFA-1 was demonstrated by competitive inhibition using free cIBR peptide or by using the I domain of LFA-1 to inhibit the binding of cIBR-NPs. In addition, T-cell adhesion to epithelial cells was inhibited by cIBR-NPs. In chapter 4, nanoparticles capable of blocking LFA-1/ICAM-1 interaction were then studied as inhibitors of T cell conjugation to DCs. LABL-NPs and cIBR-NPs rapidly bound to DCs and inhibited T cell conjugation to DCs to a greater extent than the free peptides, unconjugated NPs, anti-ICAM-1 antibodies and anti-LFA-1 antibodies. In addition, DCs treated with NPs or with cIBR-NPs stimulated the proliferation of T cells, but DCs treated with LABL-NPs did not stimulate T cell proliferation. LABL-NPs and cIBR-NPs also altered cytokine production compared to free ligands suggesting these NPs may offer a unique tool for shaping T cell response. In chapter 5, multivalent ligands having both ovalbumin (OVA) antigen and LABL peptide grafted to hyaluronic acid (HA) were found to bind professional APCs and may offer an alternative targeting approach for inducing immune tolerance. Collectively, results verified that cyclic and linear LABL and cIBR peptides can target NPstoICAM-1 and LFA-1, respectively, to deliver encapsulated agent or to provide function as potent immune modulators.